The long term goal of this investigation is to understand the mechanisms regulating the spatial and temporal aspects of collagen mineralization. During the last grant period, partial characterization of the chemical states of collagen cross-linking and the three dimensional structure of the collagen fibrils in relation to mineralization were accomplished. These findings have led to the hypothesis that, in collagen-based mineralized tissues, the orientation and stacking of mineral platelets are spatially regulated by the configuration of the nontriple helical domains (telopeptides) of the type I collagen molecules. In addition, a group of proteoglycans (many of them are collagen-associated proteoglycans) in the tooth have been identified and were found to be immunolocalized almost exclusively in the premineralizing matrices such as near pericementocytes, precementum and predentin. These proteoglycans may be inhibiting premature mineralization through their association with collagen fibrils. By using the tooth (which does not remodel) as the major model, the specific questions asked in this application are: What are the chemical states of the amino- and carboxyl-terminal nontriple helical domains in premineralizing, mineralizing and non (never)-mineralizing collagens? How do these chemical states change with mineralization? What are the structural characteristics of the proteoglycans and their spatial relationship to collagen fibrils in predentin and precementum? and how do they change in mineralizing matrix (dentin and cementum)? In order to address these questions, biochemical, physical, immunochemical and immunohistochemical techniques will be employed. The information obtained from these studies will provide insight into the regulatory mechanisms of collagen mineralization.